Lyme Disease is the most common vector-borne illness in the U.S. and is caused by transmission of the spirochete, Borrelia burgdorferi, via the tick Ixodes scapularis. Among the clinical manifestations of Lyme Disease is arthritis, which can become chronic and resistant to antibiotics. A significant proportion of the lymphocytes that accumulate in Lyme arthritis synovial fluid are gamma/delta (??? T cells of the Vd1 subset. We have determined that the synovial V41 cells are highly responsive to B. burgdorferi lipopeptides (that bind Toll-like Receptor 2, TLR2), express high levels of Fas-Ligand (FasL), may react to CD1b, and resemble chronically activated T cells. Furthermore, the Vd1 cells stimulate effector function of dendritic cells (DC) via FasL, which then feeds back to activate the Vd1 cells. DC are resistant to FasL-mediated cell death due to high level expression of the Fas death receptor inhibitor, c-FLIP. c-FLIP diverts signals from the caspase cascade and toward the NF-?B pathway. Thus, Fas/FasL may be an important link between ?? T cells and activation of DC. Mice lacking functional FasL manifest reduced inflammation and arthritis with B. burgdorferi infection. This project thus examines three closely related components of Lyme arthritis: B. burgdorferi, synovial Vd1 T cells, and dendritic cells (DC). The model connecting these three components is that B. burgdorferi binds TLR on DC, which signal upregulation of molecules such as CD1b that are stimulatory for synovial Vd1 cells. This will be also studied by in vivo infection with B. burgdorferi in mice lacking TLR, MyD88, or CD1d. A soluble Vd1 TCR will also be made to identify other Vd1 ligands (Aim 1). The synovial Vd1 cells become repeatedly activated by CD1b and Borrelia lipopeptides, as well as by properties intrinsic to their TCR/CD3 composition, which results in high level expression of surface Fas-ligand (FasL) (Aim 2). DC in turn receive stimulatory signals also via FasL from V41 cells. In the presence of high levels of the Fas inhibitor FLIP in DC, Fas signals are diverted from death pathways to growth and differentiation signals via NF-?B. This occurs via recruitment to c-FLIP of the adaptor proteins, RIP1 and TRAF2. TLR signaling of DC may also require caspase-8 and connect via TRAF6. Finally, the in vivo role of Fas/FasL will be studied by adoptive transfer of FasL+ ?? T cells to mice bearing mutant FasL or lacking Fas, caspase-8, or c-FLIP selectively in macrophages and/or DC (Aim 3). This work represents the only known research on human ?? T cells in Lyme arthritis. Project Narrative ?? T cells remain an enigma in the immune system. They are often localized at epithelial barriers, and are felt to be involved in the initial response to various infections. Indeed a protective role of ?? T cells has been observed in various infectious models. ?? T cells also accumulate at sites of inflammation in autoimmune syndromes such as in the synovial tissue in rheumatoid arthritis, the bowel in celiac disease, or the lungs in sarcoidosis. However, almost nothing is nothing regarding the specificity of ?? T cells. We are thus using Lyme arthritis in humans and mice as a model of the ?? T cell response in infection and potentially an autoimmune situation, as patients with chronic antibiotic-resistant Lyme arthritis closely resemble autoimmune rheumatoid arthritis in the composition of the synovial tissue, response to immunosuppressive agents, and even the same HLA-DR4 association. This project will seek to establish both the specificity of synovial Vd1 cells using a soluble TCR-Vd1, as well as their effector function through high expression of Fas- ligand and their ability to stimulate dendritic cells by Fas. Understanding these interactions will improve our understanding of how ?? T cell regulate the immune response during infection and autoimmune conditions.